The proposed experiments are designed to compare and contrast the regulation and the nature of hormone secretion by canine pituitary intermediate lobe cells with cells obtained from rats. In vivo experiments conducted in our laboratory have demonstrated that the canine pituitary intermediate lobe is likely regulated in a different manner than the rat lobe. Further, while the rat intermediate lobe consists of a monotonous cell population that secretes Alpha-MSH, corticotropin-like intermediate peptide, and Beta-endorphin-like materials, two distinct cell types have been identified using immunocytochemical staining in the dog lobe. The predominant cell type in the canine intermediate lobe appears identical to cells in the rat lobe, while the second cell type stains for ACTH and Beta-lipotropin in a fashion identical to pars distalis corticotrophs. These previous studies raise the question of whether canine pituitary intermediate lobe cells secrete ACTH under physiologic or pathologic conditions. To examine pituitary intermediate lobe regulation and secretion, neurointermediate lobes will be collected from healthy dogs and rats immediately after euthanasia. The lobes will be diced and the fragments incubated with trypsin to dissociate the cells. The cells will then be absorbed onto an inert matrix and inserted into a continuous perifusion system. Various neuropharmacologic agonists (acetylcholine, dopamine, norepinephrine, epinephrine, serotonin), antoagonists (atropine, haloperidol, phenoxybenzamine, propranolol, cyproheptadine), and known ACTH regulators (corticotropin-releasing factor, glucocorticoids) will be infused alone and in certain combinations. The effluent from the cells will be collected and assayed for ACTH, Alpha-MSH, and in some instances Beta-endorphin/Beta-lipotropin content by radioimmunoassay. Since canine Cushing's syndrome serves as a model for the human disease, results from these experiments may later be applied to clarify the possible contribution of intermediate lobe cells to pituitary dysfunction and also to help demonstrate the potential for subcategories of pituitary-dependent disease. Future studies would involve perifusion of adenomatous pituitary intermediate lobe tissue obtained from dogs with Cushing's syndrome, examination of the potential regulatory role of other bioactive substances on intermediate lobe cells, and an attempt to separate intermediate lobe ACTH-secreting cells from Alpha-MSH-secreting cells and to compare their regulation with pars distalis corticotrophs.